RESUMO
OBJECTIVE: To reveal specific gene activation in nitric oxide (NO)-related inflammation we studied differential gene expression in inflammatory bowel disease (IBD). METHODS: Total RNA was isolated from 20 biopsies of inflamed mucosa from Crohn's disease (CD) and ulcerative colitis (UC) patients each as well as from six controls, labeled with (32)P-dCTP and hybridized to a human NO gene array. Significant genes were analyzed for functional gene interactions and heatmaps generated by hierarchical clustering. A selection of differentially expressed genes was further evaluated with immunohistochemical staining. RESULTS: Significant gene expression differences were found for 19 genes in CD and 23 genes in UC compared to controls, both diseases with high expression of ICAM1 and IL-8. Correlation between microarray expression and corresponding protein expression was significant (r = 0.47, p = 0.002). Clustering analysis together with functional gene interaction analysis revealed clusters of coregulation and coexpression in CD and UC: transcripts involved in angiogenesis, inflammatory response mediated by the transcription factor hypoxia-inducible factor 1, and tissue fibrosis. Also, a fourth cluster with transcripts regulated by the transcription factor Sp1 was found in UC. CONCLUSIONS: Expression analysis in CD and UC revealed disease-specific regulation of NO-related genes, which might be involved in perpetuating inflammatory disease activity in IBD.
Assuntos
Colite Ulcerativa/genética , Doença de Crohn/genética , Expressão Gênica , Óxido Nítrico/metabolismo , Transdução de Sinais/genética , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Estudos de Casos e Controles , Distribuição de Qui-Quadrado , Análise por Conglomerados , Colite Ulcerativa/metabolismo , Colite Ulcerativa/patologia , Doença de Crohn/metabolismo , Doença de Crohn/patologia , Feminino , Fibrose/genética , Perfilação da Expressão Gênica , Humanos , Fator 1 Induzível por Hipóxia/genética , Molécula 1 de Adesão Intercelular/genética , Molécula 1 de Adesão Intercelular/metabolismo , Interleucina-1beta/genética , Interleucina-1beta/metabolismo , Interleucina-8/genética , Interleucina-8/metabolismo , Masculino , Metaloproteinase 1 da Matriz/genética , Metaloproteinase 1 da Matriz/metabolismo , Pessoa de Meia-Idade , NF-kappa B/genética , NF-kappa B/metabolismo , Neovascularização Patológica/genética , Óxido Nítrico/genética , Óxido Nítrico Sintase Tipo II/genética , Óxido Nítrico Sintase Tipo II/metabolismo , Análise de Sequência com Séries de Oligonucleotídeos , Fator de Transcrição Sp1/genética , Estatísticas não Paramétricas , Adulto JovemRESUMO
AIM: To explore rectal nitric oxide (NO) as biomarker of treatment response in ulcerative colitis (UC) and Crohn's disease (CD), and examine relationships between rectal NO, mucosal expression of NO synthases (NOS), and pro-inflammatory cytokines. METHODS: Twenty-two patients with UC and 24 with CD were monitored during steroid treatment. Rectal NO levels were measured and clinical activities were assessed on days 1, 3, 7 and 28. Mucosal presence of NOS and pro-inflammatory cytokines were analyzed by immunohistochemistry and RT-PCR. RESULTS: Active UC and CD displayed markedly increased rectal NO levels (10950 +/- 7610 and 5040 +/- 1280 parts per billion (ppb), respectively) as compared with the controls (154 +/- 71 ppb, P < 0.001). Rectal NO correlated weakly with disease activity in both UC and CD (r = 0.34 for UC and r = 0.48 for CD, P < 0.01). In 12 patients, a steroid-refractory course led to colectomy. These patients had only slightly increased NO levels (UC: 620 +/- 270 ppb; CD: 1260 +/- 550 ppb) compared to those with a therapeutic response (UC: 18860 +/- 530 ppb, P < 0.001; CD: 10060 +/- 3200 ppb, P < 0.05). CONCLUSION: Rectal NO level is a useful biomarker of treatment response in IBD as low NO levels predicts a poor clinical response to steroid treatment.
Assuntos
Colite Ulcerativa/fisiopatologia , Doença de Crohn/fisiopatologia , Óxido Nítrico/análise , Reto/química , Adolescente , Adulto , Idoso , Biomarcadores/análise , Colite Ulcerativa/tratamento farmacológico , Colite Ulcerativa/patologia , Doença de Crohn/tratamento farmacológico , Doença de Crohn/patologia , Resistência a Medicamentos , Feminino , Glucocorticoides/uso terapêutico , Humanos , Imuno-Histoquímica , Interferon gama/análise , Interferon gama/fisiologia , Interleucina-1/análise , Interleucina-1/fisiologia , Mucosa Intestinal/química , Mucosa Intestinal/patologia , Mucosa Intestinal/fisiopatologia , Masculino , Pessoa de Meia-Idade , Neurônios/enzimologia , Óxido Nítrico/fisiologia , Óxido Nítrico Sintase Tipo I/análise , Óxido Nítrico Sintase Tipo I/fisiologia , Óxido Nítrico Sintase Tipo II/análise , Óxido Nítrico Sintase Tipo II/fisiologia , Óxido Nítrico Sintase Tipo III/análise , Óxido Nítrico Sintase Tipo III/fisiologia , Valor Preditivo dos Testes , Resultado do Tratamento , Fator de Necrose Tumoral alfa/análise , Fator de Necrose Tumoral alfa/fisiologiaRESUMO
The monomeric G-protein, Rhes, is a candidate imidazoline-regulated molecule involved in mediating the insulin secretory response to efaroxan [S.L. Chan, L.K. Monks, H. Gao, P. Deaville, N.G. Morgan, Identification of the monomeric G-protein, Rhes, as an efaroxan-regulated protein in the pancreatic beta-cell, Br. J. Pharmacol. 136 (1) (2002) 31-36]. This suggestion was based on observations regarding changes in Rhes mRNA expression in rat islets and pancreatic beta-cells after prolonged culture with efaroxan, leading to desensitization of the insulin response to the compound. To verify this report, we have evaluated the effects of the imidazoline compounds efaroxan and BL11282 on Rhes mRNA expression in isolated rat pancreatic islets maintained in conditions identical to those used by Chan et al. The results demonstrate that desensitization of the insulin response to efaroxan, or to another imidazoline, BL11282, does not change Rhes mRNA expression levels. Transfection of MIN6 cells with plasmids containing Rhes or Rhes-antisense also does not alter efaroxan- or BL11282-induced insulin secretion. Together, these data do not support the hypothesis that Rhes is an imidazoline-regulated protein.